All procedures involving human participants have been performed in compliance with the institutional, national and international guidelines for human welfare and have been reviewed by the local institutional review board.

1. Staining 
<ol>
	<li>Before proceeding with staining, wipe off any excess solutions around the tissue on the slide with tissue paper.</li>
	<li>Place the slides in room temperature and room humidity conditions to avoid drying of the tissue on the slide throughout the entire staining process.</li>
	<li>Oxidize the slides with a few drops of potassium permanganate for 5 min. Ensure that the volume of potassium permanganate is enough to cover the tissue section on each slide.</li>
	<li>Rinse the oxidized slides with distilled water in a Coplin jar with 2-3 changes. Observe the blue color of the tissue.</li>
	<li>Now, bleach these slides by adding a few drops of oxalic acid for 5 min. Ensure that the volume of oxalic acid is enough to completely cover the tissue section on each slide.</li>
	<li>Rinse the slides by immersing them in the Coplin jar filled with distilled water. Perform the rinse 2x-3x.</li>
	<li>Now, wash these slides briefly in 95% alcohol, and then immerse the slides into an elastin solution (5 g/L) for 8-24 h. 
	NOTE: Elastin solution is volatile, and its staining time is relatively long, so it is best to immerse the slides with elastic staining in a sealed container, such as a transparent glass bottle with a sealing cap. The capacity of the container and the volume of the elastin solution depends on the number of immersed slides, as long as it enables the complete immersion of all the slides.</li>
	<li>Immerse these slides directly in 95% ethanol for 1-2 min to differentiate each tissue section well. 
	NOTE: Differentiation refers to the change in the charge of the tissue section, which removes the excess staining absorbed by the tissue and makes the colors clearer. After elastin solution staining, samples should not be washed with distilled water to avoid any difficulty in differentiation. Check microscopically, as necessary, for blue-black elastic fibers staining and gray background. It is better to slightly under differentiate the tissue since the subsequent Van Gieson's counterstain can also somewhat extract the elastic stain.</li>
	<li>Fully rinse these slides with distilled water after differentiation.</li>
	<li>Counterstain the slides in Van Gieson's solution for 1 min. 
	NOTE: The volume of Van Gieson's solution needs to be enough to be able to immerse the tissue section on each slide completely.</li>
</ol>

<table><tbody><tr><td>Elastin-Van Gieson staining kit</td><td>Baso</td><td>BA4083B</td><td>Used for staining of elastic fibers, collagen fibers and myofibers</td></tr><tr><td>Ethanol</td><td>LMAI Bio</td><td>LM64-17-5</td><td>100%</td></tr></tbody></table>

elastic fiber staining of slide sections: a technique to observe gastric cancer metastasis

Elastic Fiber Staining of Slide Sections: A Technique to Observe Gastric Cancer Metastasis

Source: Lei, G. et al. <a href='https://www.jove.com/video/58278' target='_blank'>Elastic Staining on Paraffin-embedded Slides of pT3N0M0 Gastric Cancer Tissue.</a> J. Vis. Exp. (2019)
In this video, we demonstrate the elastic staining of sections obtained from gastrointestinal tumor tissue. This staining procedure enables visualization of fine elastic fibers and helps determine tumor cell invasion beyond the elastic lamina.

Source: Lei, G. et al. Elastic Staining on Paraffin-embedded Slides of pT3N0M0 Gastric Cancer Tissue. J. Vis. Exp. (2019)
In this video, we demonstrate ...

Elastic fiber staining helps examine cancer cell invasion beyond the peritoneal elastic lamina or the sub-mesothelial elastic tissue.
To begin, add potassium permanganate solution to a processed tissue section, followed by immersion in oxalic acid. The strong oxidizing property of the potassium permanganate-oxalic acid combination bleaches the natural pigment from the tissue section to prevent interference with the staining process by obscuring elastic fiber morphology.
Rinse the slide with water and ethanol to remove any residual bleaching agent. Next, incubate the specimen in an elastin stain that comprises an iron-hematoxylin complex. Elastin, the main elastic fiber component, has a strong affinity for iron-hematoxylin complexes.&#160;
Wash the slide with ethanol and water to remove any residual stain. Finally, treat the specimen with a counterstain containing picric acid and Ponceau S. The smaller picric acid molecules penetrate all tissue types but are firmly retained by the close textured muscle and cancer cells. In contrast, the larger Ponceau S molecules displace picric acid in the porous-textured collagen, causing collagen to be stained differently from the muscle and cancer cells.
When observed under a microscope, elastic fibers appear as blue-black filaments. Collagen stains red, while muscle and cancer cells appear yellow. Yellow cancer cells penetrating the blue-black elastic lamina confirm metastatic invasion.

First, use tissue paper to wipe off any excess solution on the slide or around the tissue. Place the slides in conditions that match room temperature and humidity. Add a few drops of potassium permanganate to each slide to oxidize them, making sure that the volume added is enough to cover the tissue section on each slide. After 5 minutes, rinse the oxidized slides with distilled water in Coplin jars, repeating the rinse two to three times, and using a new jar of water for each rinse. The tissue should have an observable brown color.
Now, add a few drops of oxalic acid to each slide to bleach them, making sure that the volume added is enough to cover the tissue section on each slide. After 5 minutes, rinse the bleach slides with distilled water, repeating the rinse two to three times, and using a new jar of water for each rinse. Wash the slides briefly in 95% alcohol.
Immerse the washed slides in elastin solution for 8 to 24 hours. Then, immerse the slides directly in 95% ethanol for 1 to 2 minutes to differentiate each tissue section well. Rinse the slides completely with distilled water.
Check microscopically for the blue-black elastic fibers staining and the gray background. It's better to slightly under differentiate the tissue since the subsequent Van Gieson's counterstain can also somewhat extract the elastic stain.
Counterstain the slides in a Van Gieson solution for 1 minute, making sure that the volume used is enough to cover the tissue section on each slide completely. Next, rapidly drop 95% ethanol onto the tissue slices for a few seconds to differentiate each tissue section well.

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2.7K Views. Elastic fiber staining helps examine cancer cell invasion beyond the peritoneal elastic lamina or the sub-mesothelial elastic tissue. To begin, add potassium permanganate solution to a processed tissue section, followed by immersion in oxalic acid. The strong oxidizing property of the potassium permanganate-oxalic acid combination bleaches the natural pigment from the tissue section to prevent interference with the staining process by obscuring elastic fiber morphology. Rinse th...

Elastic Fiber Staining of Slide Sections: A Technique to Observe Gastric Cancer Metastasis

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Tags

Name	Company	Catalog Number	Comments
Elastin-Van Gieson staining kit	Baso	BA4083B	Used for staining of elastic fibers, collagen fibers and myofibers
Ethanol	LMAI Bio	LM64-17-5	100%